Quantification of the expression levels of lysosomal cysteine proteinases in purified human osteoclastic cells by competitive RT-PCR

Calcified Tissue International
O IshibashiM Kumegawa

Abstract

Cathepsin K is a lysosomal cysteine proteinase (LCP) predominantly expressed in osteoclasts. This study was conducted to evaluate the importance of human cathepsin K for osteoclastic bone resorption relative to that of other LCPs. To accomplish this, we quantitatively determined the expression levels of major LCPs (cathepsins B, K, L, and S) in human osteoclastic cells by using competitive RT-PCR. Giant cell tumor of bone (GCT) was used as a source of human osteoclastic cells, since the tissue was shown to contain a large number of cells satisfying the criteria for typical osteoclasts. The involvement of LCPs in the bone-resorption process by the GCT cells was confirmed by showing that trans-epoxysucciny-L-leucylamido-(4-guanidino) butane (E-64), a nonselective cysteine proteinase inhibitor, exerted an inhibitory effect on the pit formation. We isolated osteoclast-like cells (OLCs) positive for tartrate-resistant acid phosphatase (TRAP) and cathepsin K from the GCT tissue to a degree of almost 95% purity. In these cells, the expression of cathepsin K was shown to be approximately 20-, 130-, and 410-fold stronger than that of cathepsins B, L, and S, respectively. A similar result was obtained when human bone marrow cells in cult...Continue Reading

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Citations

Jun 3, 1999·Bioorganic & Medicinal Chemistry·R W MarquisD F Veber
Apr 8, 2010·Biological Chemistry·Gaetano LetoFrancesca Maria Tumminello
Jul 25, 2003·Microscopy Research and Technique·Jean-Marie DelaisséLaurence Blavier
Feb 26, 2004·Neurourology and Urodynamics·Bertha ChenMary Lake Polan
Dec 3, 2005·Matrix Biology : Journal of the International Society for Matrix Biology·Tiina Laitala-LeinonenAri Rinne
Jul 28, 2004·The American Journal of Pathology·Jan H N LindemanPancras C W Hogendoorn
Feb 3, 2018·Histochemistry and Cell Biology·Takehito Ono, Tomoki Nakashima
Sep 9, 2018·Calcified Tissue International·Dinisha Cyril PirapaharanJean-Marie Delaisse
Jun 3, 2005·The Journal of Biological Chemistry·Jenny LjusbergGöran Andersson

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